Biosynthesis of opiate alkaloids

Abstract

The disclosure relates to a nucleic acid molecule isolated from a Papaver somniferum cultivar that produces the opiate alkaloid noscapine which comprises 10 genes involved in the biosynthesis of opiate alkaloids.

The invention claimed is: 
     
       1. A method to produce a  P. somniferum  plant that has altered expression of a polypeptide, comprising:
 i) mutagenizing a wild-type seed from a wild-type  P. somniferum  plant that expresses said polypeptide, thereby generating mutagenized seed; 
 ii) cultivating the mutagenized seed to produce first and subsequent generations of mutagenized plants; 
 iii) obtaining seed from the first and subsequent generations of mutagenized plants; 
 iv) determining if the plants from the seed from said first and subsequent generations of mutagenized plants have an altered nucleotide sequence, altered expression of said polypeptide or both, an altered nucleotide sequence and altered expression of said polypeptide by obtaining a nucleic acid sample from the plant of said first or subsequent generation of mutagenized plants and analysing the nucleic acid sample for presence of a nucleic acid molecule comprising a nucleotide sequence having at least 90% sequence identity to the sequence of SEQ ID NO: 7, 8, 9 or 10 and that encodes a polypeptide with opiate alkaloid biosynthetic activity; and optionally 
 v) comparing the nucleotide sequence of the nucleic acid molecule in said sample to a nucleotide sequence of a nucleic acid molecule of the wild-type plant. 
 
     
     
       2. The method according to  claim 1  wherein said nucleic acid sample obtained from the mutagenized seed or plant is analysed by a method comprising:
 i) extracting nucleic acid molecules from said first and subsequent generations of mutagenized plants; 
 ii) amplifying a part of said extracted nucleic acid molecules by a polymerase chain reaction, thereby generating amplified nucleic acid comprising a part of a nucleic acid molecule comprising a nucleotide sequence having at least 90% sequence identity to the sequence of SEQ ID NO: 7, 8, 9 or 10; 
 iii) forming a preparation comprising the amplified nucleic acid and nucleic acid extracted from wild-type plants to form heteroduplex nucleic acid; 
 iv) incubating said preparation with a single stranded nuclease that cuts at a region of heteroduplex nucleic acid to identify a mismatch in said heteroduplex; and 
 v) determining the site of the mismatch in said nucleic acid heteroduplex. 
 
     
     
       3. The method according to  claim 1 , wherein said  P. somniferum  plant has enhanced opiate alkaloid biosynthetic activity. 
     
     
       4. The method according to  claim 1 , wherein said  P. somniferum  plant has reduced or abrogated opiate alkaloid biosynthetic activity. 
     
     
       5. A  P. somniferum  plant obtained by the method of  claim 1 . 
     
     
       6. The method of  claim 1 , wherein the nucleic acid sample is analysed for the presence of a nucleic acid molecule comprising the nucleotide sequence of SEQ ID NO: 7, 8, 9 or 10.